Platelet aggregation inhibitors from Bothrops alternatus snake venom

Abstract

Snake venoms are a complex mixture of proteins and peptides that can activate/inhibit platelet aggregation.Bothrops alternatus venom include three main families: metalloproteinases (SVMPs), serinoproteinases (SVSPs)and phospholipases A2 (PLA2s), among other minor components. In this work, we used inhibitor cocktails(containing Na2-EDTA, PMSF and/or pBPB) to investigate the effect of these three families and of baltergin (a PIIISVMP) on platelet aggregation by a turbidmetric method using a microplate reader. Cocktails 1 (active SVMPs)and 2 (active PLA2s) significantly reduced aggregation induced by ristocetin and collagen and by collagen andthrombin, respectively. Cocktail 3 (active SVSPs) showed a mild activation of aggregation, indicating the contentof thrombin-like enzymes (TLEs) in this venom is low. Cocktail 4 (active minor components) displayed inhibitoryeffect with all agonists assayed (ristocetin, ADP, collagen and thrombin) but at higher IC50 values. Balterginexhibited inhibitory effect when the catalytic domain was active for ristocetin-stimulated platelet aggregationand showed a non-enzymatic mechanism of inhibition when collagen was used as agonist. It was not able todisaggregate platelet thrombus. We conclude that B. alternatus venom is a source of natural inhibitors of plateletaggregation due to the action of SVMPs and PLA2s. Other minor components such as C-type lectins likelycontribute to the antiplatelet effect. The interest in knowing the action of venom components on platelet functionlies both in the understanding of the pathophysiology of snake bite envenomation and in their biotechnologicalapplication.